Cysteine-rich domains of Muc3 intestinal mucin promote cell migration, inhibit apoptosis, and accelerate wound healing
Cysteine-rich domains of Muc3 intestinal mucin promote cell migration, inhibit apoptosis, and accelerate wound healing
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DOI：
10.1053/j.gastro.2006.09.006
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发表时间：
2006-11-01
期刊：
影响因子：
29.4
通讯作者：
Shekels, Laurie L.
中科院分区：
文献类型：
--
作者：
Ho, Samuel B.;Dvorak, Leah A.;Shekels, Laurie L.
背景与目的：Muc 3肠粘蛋白含有一个富含半胱氨酸的胞外结构域，具有两个表皮生长因子（EGF）样基序。本研究的目的是确定Muc 3蛋白的功能特性。研究方法：从大肠杆菌中纯化含有Muc 3 EGF样结构域（m3 EGF 1，2）或截短形式（m3 EGF 1和m3 EGF 2）的Glucose S-转移酶融合蛋白。检查小鼠结肠（年轻成年小鼠结肠）和人A431和LoVo细胞响应重组蛋白的迁移和酪氨酸磷酸化。用人MUC 3A跨膜-EGF 1，2构建体转染LoVo细胞，并分离稳定克隆（LhM 3c 14）。特异性小干扰RNA转染可抑制LoVo中内源性MUC 3A的表达。通过细胞核形态学或末端脱氧核苷酸转移酶介导的脱氧尿苷三磷酸生物素缺口末端标记法定量细胞凋亡。通过口服5%葡聚糖硫酸钠或直肠5%乙酸诱导小鼠结肠炎，然后进行灌肠治疗。结果：m3 EGF 1，2在所有细胞系中刺激细胞迁移，但不诱导增殖。酪氨酸磷酸化抑制剂，染料木黄酮，但不是由表皮生长因子受体抑制剂，tyrphostin（AG 1478）抑制迁移。在LoVo中抑制内源性MUC 3A减少基线迁移。ErbB受体的酪氨酸磷酸化没有观察到治疗后的细胞与m3 EGF 1，2。用m3 EGF 1，2预处理的LoVo细胞和转染的LhM 3c 14细胞显示对肿瘤坏死因子α或Fas受体刺激的响应的凋亡减少。经直肠给予m3 EGF 1，2可显著减少实验性急性结肠炎的粘膜溃疡和细胞凋亡。截短蛋白m3 EGF 1和m3 EGF 2没有影响。结论：Muc 3粘蛋白富含半胱氨酸的结构域在上皮修复中起着积极的作用，并且代表了用于肠创伤愈合的潜在的新型治疗剂。
Background & Aims: Muc3 intestinal mucin contains an extracellular cysteine-rich domain with 2 epidermal growth factor (EGF)-like motifs. The aim of this study was to determine the functional properties of Muc3 proteins. Methods: Glutathione S-transferase-fusion proteins containing both Muc3 EGF-like domains (m3EGF1,2) or truncated versions (m3EGF1 and m3EGF2) were purified from Escherichia coli. Mouse colon (young adult mouse colon) and human A431 and LoVo cells were examined for migration and tyrosine phosphorylation in response to recombinant proteins. LoVo cells were transfected with a human MUC3A transmembrane-EGF 1,2 construct and a stable clone was isolated (LhM3c14). Endogenous MUC3A in LoVo was inhibited by specific small interfering RNA transfection. Apoptosis was quantitated by nuclear morphology or terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate biotin nick-end labeling assay. Colitis was induced in mice by oral 5% dextran sodium Sulfate or rectal 5% acetic acid, followed by enema treatments. Results: m3EGF1,2 Stimulated cell migration in all cell lines, but did not induce proliferation. Migration was inhibited by a tyrosine phosphorylation inhibitor, genistein, but not by the EGF receptor inhibitor, tyrphostin (AG1478). Inhibition of endogenous MUC3A in LoVo reduced baseline migration. Tyrosine phosphorylation of ErbB receptors was not observed after treatment of cells with m3EGF1,2. LoVo cells pretreated with m3EGF1,2 and transfected LhM3c14 cells showed reduced apoptosis in response to tumor necrosis factor alpha or Fas-receptor stimulation. Administration of m3EGF1,2 per rectum significantly reduced mucosal ulceration and apoptosis in experimental acute colitis. Truncated proteins m3EGF1 and m3EGF2 had no effect. Conclusions: The Muc3 mucin cysteine-rich domain plays an active role in epithelial restitution, and represents a potential novel therapeutic agent for intestinal wound healing.